NADP‐dependent malate dehydrogenase (decarboxylating) from sugar cane leaves

Abstract

NADP-dependent malate dehydrogenase (decarboxylating) from sugar cane leaves was inhibited by increasing the ionic strength in the assay medium. The inhibitory effect was higher at pH 7.0 than 8.0, with median inhibitory concentrations (ICs0) of 89 mM and 160 mM respectively, for inhibition by NaCl. Gel-filtration experiments indicated that the enzyme dissociated into dimers and monomers when exposed to high ionic strength (0.3 M NaCl). By using the enzyme-dilution approach in the absence and presence of 0.3 M NaC1, the kinetic properties of each oligomeric species of the protein was determined at pH 7.0 and 8.0. Tetrameric, dimeric and monomeric structures were shown to be active but with different V and K , values. The catalytic efficiency of the oligomers was tetramer > dimer > monomer, and each quaternary structure exhibited higher activity at pH 8.0 than 7.0.Dissociation constants for the equilibria between the different oligomeric forms of the enzyme were determined. It was established that Kd values were affected by pH and Mg2+ levels in the medium. Results suggest that the distinct catalytic properties of the different oligomeric forms of NADP-dependent malate dehydrogenase and changes in their equilibrium could be the molecular basis for an efficient physiological regulation of the decarboxylation step of C4 metabolism.In some C4 plants such as maize, sugar cane and sorghum, the decarboxylation step of the C4 metabolic pathway of photosynthesis occurs in the chloroplast of bundle-sheath cells with oxidative decarboxylation of L-malate [I]. The reaction that takes place, L-malate + NADP+ gpyruvate + C 0 2 + NADPH, is catalysed by NADP-dependent malate dehydrogenase (oxaloacetate-decarboxylating) [I].The primary structure of maize NADP-dependent malate dehydrogenase has been recently determined [2]. Studies with the enzyme highly purified from maize [2 -51 and sugar cane [6] chloroplasts showed a homotetrameric structure of about 220 -240 kDa. However, differences have been reported concerning the quaternary subunit composition of this protein from maize, depending on the presence of dithiothreitol as well as the buffer used [7]. Recently [8], the existence of an equilibrium has been reported between dimeric and tetrameric structures in highly purified sugar cane NADP-dependent malate dehydrogenase, with pH being a potent effector of the dimer/tetramer equilibrium. Thus, it was found that at pH 7.0 or 8.0 the enzyme exists predominantly in the dimeric or tetrameric form, respectively [8].It was also reported [8] that the enzyme exhibits distinctive kinetic properties at pH 7.0 or 8.0. At pH 8.0, the enzyme is more active than at pH 7.0. It was speculated [8] physiologically important for the regulation of NADP-dependent malate dehydrogenase activity. However, a direct correlationship between quaternary structure and enzyme activity was not determined. The present paper reports kinetic studies carried out under different conditions using the enzyme-dilution technique of Kurganov [9]. The object of these s...

Molecular Basis of Aluminium Toxicity in Plants: A Review

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Evidence for the Existence of Two Essential and Proximal Cysteinyl Residues in NADP-Malic Enzyme from Maize Leaves

“…This may suggest a possible interaction of these reagents with the L-malate-binding site of the enzyme, about which very little is known (7). (12,18) seems to have no effect on the modification by maleimides. The kinetics of inactivation of the enzyme by the bifunctional maleimides OPBM, MPBM, and PPBM were biphasic (Figs.…”

Interaction of analogues of substrate with NADP-malic enzyme from maize leaves

“…NADP-ME purified from Zea mays L. (Asami et al 1979b(Asami et al , H~iusler et al 1987) and sugarcane (Iglesias and Andreo 1990a) exists as a tetrameric protein composed of identical 62kDa subunits (Rothermel and Nelson 1989). Depending on the conditions, i.e.…”

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